1887

Abstract

The gene is transcribed from four known and differently regulated promoters: P1, P3, P4 and P5. This study demonstrates that the conserved consensus sequence of the promoter in the regulatory region of the gene, described previously, is a functional promoter, P6. The evidence for this conclusion is: (i) the specific binding of the –RNAP holoenzyme to P6, (ii) the location of the transcription start site at the predicted position (C, 30 nt upstream of ATG) and (iii) the dependence of transcription on and on an ATP-dependent activator. Nitrogen starvation, heat shock, ethanol and CCCP treatment did not activate transcription from P6 under the conditions examined. Two activators of promoters, PspF and NtrC, were tested but neither of them acted specifically. Therefore, PspFΔHTH, a derivative of PspF, devoid of DNA binding capability but retaining its ATPase activity, was used for transcription , taking advantage of the relaxed specificity of ATP-dependent activators acting in solution. In experiments overexpression of PspFΔHTH from a plasmid was employed. Thus, the -dependent transcription capability of the P6 promoter was demonstrated both and , although the specific conditions inducing initiation of the transcription remain to be elucidated. The results clearly indicate that the closed –RNAP–promoter initiation complex was formed and and needed only an ATP-dependent activator to start transcription.

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2020-04-02
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